Although viral vectors may deliver a transgene to liver cells with high efficiency, they also induce serious immune or inflammatory response, which precludes the practice of repeated injection. Non-viral vectors, however, although unable to produce ultra-high levels of gene expression in the liver, have the advantage of not inducing any inflammatory reactions. We have recently developed an emulsion system resembling human chylomicron remnants with hydrophobic DNA/cationic lipid complex solubilized in the interior oil phase of the emulsion. Intraportal injection of the vector into mice results in approximately 10% of liver cells (mostly hepatocytes) expressing the transgene. Although the transgene expression lasts only about one week, a second injection at day 7 fully restores expression to initial levels. This reconstituted chylomicron remnant (RCR) vector will serve as a prototype for the development of future vectors. The aims of this project are as follows: (1). To develop intravenously injectable, liver-specific vectors by conjugating hepatocyte-specific, galactose ligands to the surface of the RCR. Special emphasis will be the use of tri-antennial galactosyl moiety and ligands to the surface of the RCR. Special emphasis will be the use of tri-antennial galactosyl moiety and asialoglycoproteins linked to a lipid as a targeting ligand. (2) To construct hepatocyte-specific expression plasmids with prolonged transgene expression activity. We will use an expression plasmid containing the regulatory sequence of Adeno Associated Virus (AAV) and liver specific promoter for persistent gene expression. Inclusion of an intron and a Kozak sequence at the 5' end of the transgene coding sequence is expected is expected to further enhance the transgene expression level. 93). To demonstrate the gene therapy potential of the new vector, we will combine the AAV expression system and the RCR emulsion to construct a hybrid vector containing the mouse ornithine transcarbamylase (OTC) cDNA. The hybrid vector will be injected into the sparse fur mice, which contain mutations in the OTC gene and display only low level of OTC activity. These studies will hopefully lead to the development of a safe and effective gene therapy vector for treating human metabolic diseases such as OTC deficiency.